The goal of this investigation is to determine targets in the aflatoxin B1 biosynthetic pathway at which specific chemicals, either natural or synthetic, can be used to inhibit pathway function. One objective is to understand the enzymatic activity of the protein products 4 genes which have been cloned from the aflatoxin gene cluster. The ability to measure the conversion of substrate to end product by these enzymes depends on an accurate identification of the chemical structures involved. For example, we have used mass spectrometry together with other analytical methods including, NMR, UV/VIS spectroscopy to confirm that the protein product of the nor-1 gene is a reductase which converts norsolorinic acid to averantin. An understanding of enzyme activity may provide us with useful information allowing generation of specific inhibitors to help prevent the accumulation of this biologically active mycotoxin in food and feed.